This invention relates generally to liquid chromatography, and more specifically to a solvent supply system for use in high performance column liquid chromatography (HPLC), in which the control of the proportioning of solvents on the low pressure or inlet side of the pump is by means of a specially designed gradient suction cam.
Chromatography is a separation method in which a mixture of components (called the "sample" or "sample mixture") is placed as a zone at one end of a system containing both a stationary phase and a mobile phase. Each component of the sample distributes itself in dynamic equilibrium between the two phases in a ratio characteristic of that component. As a result, the flowing mobile phase causes each individual component zone to migrate at a characteristic rate, and the zones become separated after a period of time. In liquid absorption chromatography, the stationary phase consists of a tubular column packed with an absorbent material. The mobile phase for carrying an analysis sample through the column commonly referred to as the carrier, is a solvent mixture comprising two or more miscible liquids, which are introduced into the column. An equilibrium is established for the individual components of a sample mixture according to the "attraction" of each to the stationary phase and according to the solubility of each in the carrier solvent. The rate at which a solute passes through the column chromatograph is dependent upon the existing equilibrium points. Separations occur where the distributions differ.
All liquid chromatography systems include a moving solvent, a means for producing solvent motion such as gravity or a pump, a means for sample introduction, and a fractionating column. Operation of a liquid chromatography system with a carrier of two or more solvents mixed in constant, nonvarying proportions is referred to as isocratic operation.
It is often desirable to operate the liquid chromatographic system using a carrier in which the ratios of the liquid in the solvent mixture vary over time in accordance with some predetermined gradient. This type of operation is referred to as gradient elution, and the gradient profiles referred to as solvent programs. Within the category of gradient elution operation, the ratios in the solvent mixture can be made to increase at a fixed rate, i.e. linear gradient or at an increasing rate of change, i.e., convex gradient or at a decreasing rate of change, concave gradient by appropriate control of the solvent mixing apparatus.
There are various types of chromatography, e.g., liquid chromatography, gas chromatography, thin layer chromatography, etc. The major differences between these various chromatographic methods lie in the physical state of the mobile phase (gas or liquid), and the manner in which the stationary phase is supported, e.g., coated on an inert granular material packed in a tube, coated on an inner wall surface, etc. In all chromatographic methods, the separation objective is essentially the same, that is, distribution of the sample components between a mobile phase and a stationary phase. When the method is used for chemical analysis, a detector is commonly placed at the far end of the system to monitor the passage of the component zones as they emerge from the system. The signal from the detector is displayed on a recording device such as a strip chart recorder, and a record indicates both qualitative and quantitative information regarding the components of the sample.
It is often desirable for a chromatographic system to be able to provide high resolution (i.e., a large degree of component separation with narrow zones), evenly spaced component zones, rapid separation, and a satisfactory record from a very small sample. The behavior of the system described in these terms may be called the "performance" of the system. It is well known in the chromatographic art to improve system performance by changing one of the system variables during the course of the analysis such as temperature, chemical composition of the mobile phase, and the flow rate of the mobile phase.
An essential objective relevant to all liquid chromatography apparatus of the type considered herein, is that of providing a proper flow of solvent to and through the chromatographic column. Thus, in the past, numerous and varied approaches have been utilized for supplying solvents to high performance liquid chromatographic columns.
A key requirement in this regard is that of providing a relatively nonpulsating, constant flow of solvent. In addition, because an LC (liquid chromatography) detector is sensitive to flow rate variations, it can provide erroneous readings and exhibit excessive noise in the presence of a pulsating solvent flow. Various approaches have been utilized in the past in order to remove pulsation and other noise. In general, however, the prior art methodology was directed toward highly expensive and overly complex mechanisms for controlling pulsation. Thus, in a typical example in which a system is intended for operation in a gradient elution mode, i.e., by use of two distinct solvents, a dual cylinder pump arrangement has been utilized. Such an arrangement requires distinct cylinder pumps, including separate means for driving each of the pumps, thereby requiring separate speeds, etc.
A liquid chromatography system which utilizes a solvent pump, can control the pulsating problem by applying control means at either the low pressure or the high pressure end of the pumping stage. The low pressure end of the pumping system is the inlet or suction side of the pump. The high pressure end of the pumping means is the pumping side of the pump mechanism. The overwhelming majority of systems in the prior art are directed toward controlling pump pulsation on the high pressure end of the system.
Pulsation control has typically been provided by a complex mechanical means on the high pressure end of the system or through an electronically actuated feedback circuit which would control motor speed, or another flow parameter. Thus, in U.S. Pat. No. 4,045,343 entitled "High Pressure liquid Chromatography System", pulsation control was provided through means of a complex system of valves and control apparatus. In U.S. Pat. No. 3,985,021 entitled "High Performance Liquid Chromatography System", feedback means were provided for controlling the rotational speed of the motor throughout the reciprocating cycle of the pump so as to provide the preselected rotational speeds over predetermined subintervals of each successive reciprocation cycle. Application of the control cycle was synchronized with the pumping cycle so that the speed control was properly applied over each successive reciprocating cycle in order to control output pulsation. In U.S. Pat. No. 3,981,620 entitled "Pumping Apparatus", control on the high pressure side of the pumping mechanism was also achieved through a pressure sensing device which incorporated a feedback system to control the speed of the motor. This feedback system not only controlled the speed of the motor but provided a means to limit the current to the motor such that only the current necessary to drive the pump was provided. U.S. Pat. No. 4,245,963, entitled "Pump", disclosed a method for controlling pulsation of the output or high pressure side of the pump by means of a liquid storage device consisting of a flattened length of coiled tubing which was placed in the flow path between the two chambers to deliver flow during the low periods when the displacement elements were in reverse direction, thereby smoothing flow delivery. Finally, U.S. Pat. No. 3,981,620 also entitled "Pumping Apparatus", utilized a feedback responsive mechanism to sense the pressure of the liquid being pumped. It utilized a "flow through" meter which comprises a conduit as its pressure sensitive element.
Several prior art systems utilized mechanical analog systems incorporating specialized cam technology for control on the high pressure side of the pump. U.S. Pat. No. 4,137,011, entitled "Flow Control System For Liquid Chromatographs," provides a control system which was particularly adapted to use in multiple chamber single pump systems in which a cam driven by a speed control device such as a stepping motor is connected to a multiple chamber positive displacement piston pump arranged with its chambers and associated pumps in opposition to either other on each side of the cam. The invention also utilizes a complex feedback network which controls the speed of the pump.
The model 2010 HPLC isocratic pump by Varian Associates is an example of a current system on the market which utilizes both cam technology and an electronic feedback mechanism to control pulsation on the high pressure side of the pumping cycle. This system utilizes a concentric face cam to facilite suction and pulsation, and also incorporates a pressure feedback system for solvent compressability compensation. The system utilizes a pressure transducer which provides high resolution for accurate readout of system operating pressure. The pressure feedback system controls motor speed, based upon the actual operating back-pressure, to compensate for solvent compression and minimize pump pulsation.
While the majority of prior art systems sought to control the high pressure side of the pumping cycle, there are major advantages to be realized by the control of the low pressure or inlet side of the pump. This is particularly true where the examination of multiple solvents is desired, and there is a need to proportion the solvents evenly. In such cases, it is desirable to provide an even and nonpulsating flow of solvents from the solvent reservoirs to the pump head. The prior art systems which sought to control the high pressure side of the pumping process create a rapid unequal draw on the low pressure or inlet side of the pump. This makes the proper proportioning of multiple solvents difficult, and requires the use of expensive specialized check valves and electronic sensing means. Moreover, with the improvement in downstream pulse dampening technology, it is no longer as necessary to control pulsation through the pumping means on the high pressure side.
One system currently on the market for controlling the low pressure side of an HPLC pump is manufactured by IBM. It utilizes a cam system with three pumping cross head followers, spaced at 120.degree. intervals about the cam. While the IBM system provides constant suction on the low pressure or inlet side of the pump, it does so at the considerable expense of an additional cross-head follower, pumping head and check valve configuration. This of course, adds extra expense and complication to the pumping procedure. The pumping barrel and check valves are the most expensive parts of an HPLC pumping system.
It would be desirable to control the flow of HPLC solvent on the low pressure or inlet side of the pump by means of a two follower cross-head pumping mechanism which could provide constant suction on the inlet side of the pump by means of a specially shaped gradient cam. This would be particularly desirable in applications in which there is a need for constant suction to proportion various solvent samples. By providing constant and uniform suction, the user could get an even proportioning of solvent. Such a system would provide the user with the ability to obtain a very smooth draw of solvent on the inlet or low pressure side of the pump.
It is the purpose of the disclosed constant suction proportioning pump to provide a constant and uniform draw of solvent on the low pressure side by means of a specially shaped gradient face cam. It is a second purpose of the disclosed proportioning pump to provide a constant suction by a relatively simple and inexpensive means on the inlet side using only two cross-head followers spaced 180.degree. apart. The gradient cam of the disclosed preferred embodiment is divided unevenly by a peak and trough extending radially from the center of the cam. 170.degree. of the cam profile is used to force the piston forward and therefore pump solvent. 190.degree. of the cam profile is used to draw a constant flow of solvent on the low pressure side of the pump. Because over one half of the surface of the cam gradient is devoted toward the draw side of the pump, and the followers are spaced evenly at 180.degree. intervals, the cam provides continuous suction. The system requires no complicated software and controls any pulsation on the high pressure side with improved pulse dampening mechanisms downstream from the pumping means. The pumping means accordingly receives a steady, properly proportioned flow of solvent.